In order to gain more basic information about the regulation of ion movement across epithelial tissues, mechanisms located in the apical and basolateral cell membrane will be investigated. The processes which are involved in the movement of the chloride ion are of particular concern in this project. Cl transport plays a major role in the function of the kidney and the most potent diuretics achieve their action by inhibiting Cl transport. From the results obtained in previous experiments it was possible to construct a detailed theoretical model embracing the various mechanisms in the apical and basolateral cell membrane which are thought to be connected with Cl transport. The individual features of this model can be tested since it is known that various compounds and experimental conditions can inhibit and or stimulate selectively different transport mechanisms in the apical and basolateral cell membrane. This permits the study of individual transport processes and of the interaction between these processes. Measurements of tracer fluxes across the apithelium and across the apical cell membrane as well as intracellular ion activity determinations with ion-sensitive microelectrodes will be used to obtain specific information about mechanisms which regulate Cl, Na and K transport. Cl, Na and K transport will be studied when (1) the solution in contact with the epithelial cells is altered by ionic replacement, (2) the membrane potential is changed by voltage clamping at different potentials, (3) inhibitors such as DIDS and SITS and diuretics such as furosemide and amiloride are used. In addition, the effects on Cl transport of agents which change Na transport such as theophylline, antidiuretic hormone, prostaglandin and ouabain will be studied. Models based on network thermodynamics will be used in conjunction with the experiments. The project will be done on tight Na transporting epithelia (frog skin, urinary bladders of Necturus maculosus and Amphiuma means) mounted in chambers (1) as isolated tissue and (2) as preparations of epithelial cells without connective tissue. The main goal of the project is the achievement of a better understanding of epithelial transport processes. These processes are critical for the functioning of vital organs of the body.